Targeted genomic integration of EGFP under tubulin beta 3 class III promoter and mEos2 under tryptophan hydroxylase 2 promoter does not produce sufficient levels of reporter gene expression

Neuronal tracing is a modern technology that is based on the expression of fluorescent proteins under the control of cell type-specific promoters. However, random genomic integration of the reporter construct often leads to incorrect spatial and temporal expression of the marker protein. Targeted integration (or knock-in) of the reporter coding sequence is supposed to provide better expression control by exploiting endogenous regulatory elements. Here we describe the generation of two fluorescent reporter systems: EGFP under pan-neural marker class III β-tubulin (Tubb3) promoter and mEos2 under serotonergic neuron specific tryptophan hydroxylase 2 (Tph2) promoter. Differentiation of Tubb3-EGFP ES cells into neurons revealed that though Tubb3-positive cells express EGFP, its expression level is not sufficient for the neuronal tracing by routine fluorescent microscopy. Similarly, the expression levels of mEos2-TPH2 in differentiated ES cells was very low and could be detected only on mRNA level using PCR-based methods. Our data shows that the use of endogenous regulatory elements to control transgene expression is not always beneficial compared to random genomic integration

Comparison of American mink embryonic stem and induced pluripotent stem cell transcriptomes

BACKGROUND: Recently fibroblasts of many mammalian species have been reprogrammed to pluripotent state using overexpression of several transcription factors. This technology allows production of induced pluripotent stem (iPS) cells with properties similar to embryonic stem (ES) cells. The completeness of reprogramming process is well studied in such species as mouse and human but there is not enough data on other species. We produced American mink (Neovison vison) ES and iPS cells and compared these cells using transcriptome analysis. RESULTS: We report the generation of 10 mink ES and 22 iPS cell lines. The majority of the analyzed cell lines had normal diploid chromosome number. The only ES cell line with XX chromosome set had both X-chromosomes in active state that is characteristic of pluripotent cells. The pluripotency of ES and iPS cell lines was confirmed by formation of teratomas with cell types representing all three germ layers. Transcriptome analysis of mink embryonic fibroblasts (EF), two ES and two iPS cell lines allowed us to identify 11831 assembled contigs which were annotated. These led to a number of 6891 unique genes. Of these 3201 were differentially expressed between mink EF and ES cells. We analyzed expression levels of these genes in iPS cell lines. This allowed us to show that 80% of genes were correctly reprogrammed in iPS cells, whereas approximately 6% had an intermediate expression pattern, about 7% were not reprogrammed and about 5% had a "novel" expression pattern. We observed expression of pluripotency marker genes such as Oct4, Sox2 and Rex1 in ES and iPS cell lines with notable exception of Nanog. CONCLUSIONS: We had produced and characterized American mink ES and iPS cells. These cells were pluripotent by a number of criteria and iPS cells exhibited effective reprogramming. Interestingly, we had showed lack of Nanog expression and consider it as a species-specific feature

Incidence and prognostic value of acute kidney injury in pulmonary embolism: data from the SIRENA registry

Aim. To evaluate the incidence and severity of acute kidney injury (AKI), as well as its ability to reclassify the risk of premature mortality and association with inhospital mortality in patients with pulmonary embolism (PE) in the Russian population.Material and methods. From April 2018 to April 2019, the SIRENA Russian Multicenter Registry included patients with PE, as well as the deceased, in whom the PE was detected by autopsy. AKI was diagnosed according to current KDIGO guidelines (2012). Creatinine calculated according to the MDRD equation and corresponding to a glomerular filtration rate of 75 ml/min/1,73 m2 (baseline) was taken as the initial one, with subsequent assessment relative to the parameter value upon admission. The risk stratification of early death was carried out in accordance with the current ESC clinical guidelines (2019).Results. A total of 604 patients with PE were examined (men — 293 (49%), women — 311 (51%), mean age — 64±15 years). AKI was diagnosed in 223 (37%) of them. Stage 1 AKI was detected in 146 (65%), 2 — in 55 (25%), 3 — in 22 (10%) patients. Prior chronic kidney disease was recorded in 61 (10%) patients. Seventy-one (12%) patients had a high risk of death, 364 (61%) — intermediate risk, and 164 (27%) — low risk. The AKI incidence increased as the severity of PE increased: at low risk of death — 26%, intermediate — 38%, high — 59% (p<0,0001). In total, 107 (18%) patients died in the hospital. AKI led to an increase in mortality within following risk groups: at low risk, this effect was a trend (6 (5%) vs 6 (14%); p=0,052); at intermediate and high risk, significant differences was obtained (30 (13%) vs 41 (30%), p<0,001; 4 (14%) vs 19 (45%), p=0,006, respectively). Multivariate Cox regression demonstrated that AKI is a predictor of inhospital death (odds ratio (OR), 3,66 (95% confidence interval (CI): 2,37-5,66; p<0,0001), regardless of increased troponin levels (OR, 1,31 (95% CI: 0,80-2,14; p=0,28) and right ventricular dysfunction (OR, 1,23 (95% CI: 0,74-2,04; p=0,42).Conclusion. Thirty-seven percent of Russian patients with PE have AKI diagnosed by baseline creatinine. In 2/3 of the examined patients, stage 1 AKI is observed. The AKI incidence increases as the severity of PE increases. The presence of AKI reclassifies patients into a higher risk category for death and is associated with a significant increase in inhospital mortality

Hypertension control during the COVID-19 pandemic: results of the MMM2021 in Russia

Repetitive quarantines and social restrictions during the coronavirus disease 2019 (COVID-19) pandemic have negatively affected the population health in general, and the control of hypertension (HTN) in particular.Aim. To evaluate the control of HTN in the Russian population during the COVID-19 period based on the results of screening for HTN May Measurement Month 2021 (MMM2021).Material and methods. During May-August 2021, 2491 participants from 11 Russian regions took part in the screening. Participation was voluntary without restrictions on sex. All participants were over 18 years of age. During the screening, blood pressure (BP) was measured three times using automatic and mechanical BP monitors. In addition, a questionnaire was filled out on behavioral risk factors, comorbidities and therapy. HTN was diagnosed with systolic BP ≥140 mmHg and/ or diastolic blood pressure ≥90 mmHg and/or taking antihypertensive therapy. The questionnaire included questions about prior COVID-19, vaccinations and their impact on the intake of antihypertensive drugs.Results. The analysis included data from 2461 respondents aged 18 to 92, of which 963 were men (39,1%). The proportion of hypertensive patients was 41,0%, while among them 59,0% took antihypertensives and 30,9% were effectively treated. In comparison with pre-pandemic period according to MMM2018-2019, the higher proportion of HTN patients in the Russian sample was revealed during MMM2021 (41,0% vs 31,3%, p<0,001) with a comparable proportion of patients receiving antihypertensive therapy (60,7% vs 59,0%, p=0,05) and treatment efficacy (28,7% vs 30,9%, p=0,36). Monotherapy was received in 44,7% of cases, while dual and triple combination therapy — in 30,9% and 14,1%, respectively. The majority of respondents (~90%) did not adjust their antihypertensive therapy during the COVID-19 pandemic.Conclusion. According to HTN screening in Russia, there is persistent ineffective control of HTN, which may be due to both the worsening pattern of behavioral risk factors, limited access to healthcare during COVID-19, and the inertia of physicians and low adherence of patients due to the asymptomatic HTN course in the majority

Approaches to the therapy of heart failure with reduced ejection fraction. Resolution of an online meeting of the Volga Federal District experts

At an online meeting of experts held on May 14, 2021 additional research results on a sodium-glucose co-transporter-2 (SGLT2) inhibitor empagliflozin in patients with heart failure with reduced ejection fraction were considered. According to the data from the EMPEROR-Reduced international study, cardiovascular and renal effects of empagliflozin therapy in patients with and without type 2 diabetes (T2D) were analyzed. A number of proposals and recommendations was accepted regarding the further study of cardiovascular and renal effects of empagliflozin and its use in clinical practice in patients with heart failure, regardless of the T2D presence

COMPLICATIONS OF THROMBOLYTIC THERAPY IN ACTUAL CLINICAL PRACTICE

Reperfusion therapy, in particular, thrombolysis in acute myocardial infarction involves development of serious complications. This clinical situation demonstrates the complexity of treatment of patients with hemorrhagic complications after thrombolysis and makes questions on the application of antiplatelet agents and anticoagulants in such cases.</p

COMPLICATIONS OF THROMBOLYTIC THERAPY IN ACTUAL CLINICAL PRACTICE

Reperfusion therapy, in particular, thrombolysis in acute myocardial infarction involves development of serious complications. This clinical situation demonstrates the complexity of treatment of patients with hemorrhagic complications after thrombolysis and makes questions on the application of antiplatelet agents and anticoagulants in such cases

ACUTE KIDNEY INJURY IN PATIENTS WITH ACUTE DECOMPENSATED CHRONIC HEART FAILURE

Objective: to estimate the frequency and magnitude of acute kidney injury (AKI) in patients with acute decompensated chronic heart failure(ADCHF) and to clarify the relationship of AKI to mortality.Subjects and methods. One hundred and four patients (58 men and 46 women; mean age 65.3 ± 10.68 years) with ADCHF were examined.AKI was diagnosed and classified by the KDIGO criteria.Results. In terms of creatinine, AKI was diagnosed in 74 (71 %) patients (Stage I in 51 (49 %), Stage II in 20 (19 %), and Stage III in 3 (3 %)patients. Five (5 %) patients died during hospitalization. All the dead patients had AKI. Multivariate regression analysis demonstrated that regardless of gender, age, chronic heart failure stage, the in-hospital mortality was associated with the level of creatinine (R = 0.29; β = 0.20;p = 0.046). At the same time, in the patients with AKI Stages II-III the probability of in-hospital mortality was higher than that in the other patients (relative risk, 23.4; 95 % confidence interval 2.9–187.0; p = 0.003).Conclusion. More than half of the patients with ADCHF have AKI according to the KRIGO criteria. The in-hospital mortality is much higheramongst the patients with AKI Stages II-III.</p

RISK FACTORS OF ARRHYTHMIAS IN PATIENTS WITH ACUTE DECOMPENSATION OF CHRONIC HEART FAILURE

The aim of the research. The aim of this study was to investigate risk factors of arrhythmias, the frequency of acute kidney injury (AKI) in patients with acute decompensation of chronic heart failure (ADCHF) and AKI impact on dynamics of electrocardiographic parameters. Materials and methods.103 patients with acute decompensation of chronic heart failure who were survival on 10 hospital day were studied. Twenty-four-hour ECG recordings were performed on 1 and 10 days of hospitalization using automatic algorithm to measure QT and heart rate for analyze the arrhythmogenic factors. AKI was diagnosed according to KDIGO Guidelines (2012). Results. AKI was revealed in 25 (24,3%) patients with ADCHF. At admitted to the hospital the corrected interval QT (QTc) &gt; 440 ms had 42(41%) patients with ADCHF. Stabilization of the state is characterized by shortening of the QTc interval. In men, AKI had impact on the QTc: 475,50±31,72 ms vs. 445,16±29,67 ms without AKI (р=0,02). This effect persists until discharge from hospital. In women, the effect of AKI on the QTc prolongation was not detected. Patients with AKI had more premature ventricular complexes: 622,0 (128,0; 1488,0) premature ventricular complexes vs. 389.0 (42,0; 1237,0) premature ventricular complexes in patients without AKI on admission (p=0,005 ), 501,0 (81,0; 1150,0) premature ventricular complexes in patients with AKI vs. 325,0 (70,0; 1228,0) without AKI (p &lt;0,001) on the 10th day of treatment. In sinus rhythm revealed an increase of the index of variability of QT and pathological significance of heart rate turbulence. The values of heart rate variability, exceeding the “risk-sharing point of death” did not improve on 10 day treatment. Conclusion. At admitted to the hospital QTc prolongation was found in 42 (41%) patients with ADCHF. In men, the presence of AKI was associated with the QTc prolongation. Although there is clinical stabilization in patients with acute decompensated of chronic heart failure on 10 day treatment arrhythmogenic factors are saved